The current invention is generally directed toward a microcolumn and a method of loading a microcolumn. The microcolumn produced by the method of the invention is capable of separating an analyte from a sample in the millisecond time scale by using small column volumes that can tolerate medium to high flow rates.
One area of continuing interest in the field of analytical chemistry is the development of separation techniques that achieve extraction of an analyte from a sample in the millisecond time domain. The ability to rapidly and accurately separate analytes from a sample has a number of important applications in the health, pharmaceutical, clinical, research and environmental fields. One such application, for example, is the ability to rapidly extract the biologically active form of a drug, hormone, or toxin from a biological sample. This is of particular interest in clinical chemistry and pharmaceutical science as a means for controlling and studying the effect of drugs and/or hormones within the body.
Techniques such as microbore HPLC and capillary HPLC have attempted to decrease separation times and increase separation efficiency by decreasing column diameter with a corresponding increase in column length. The small column diameter acts to decrease column volume, while the increased column length serves to provide more contacts between the analyte and the separation medium. Although these techniques have increased separation efficiency by obtaining narrower elution peaks, they have not resulted in separations in the millisecond time domain.
In another approach, some techniques have attempted to decrease extraction time by reducing column lengths. Many HPLC-based immunoaffinity columns are now in the size range of a few millimeters to a few centimeters. However, columns with these dimensions, while separating an analyte from a sample in a few seconds, cannot achieve separation in the millisecond time domain. Additionally, membrane supports based on polymeric materials that are several millimeters to several centimeters in diameter and several millimeters in length have been developed. However, like the immunoaffinity columns previously discussed, the membrane supports are not capable of separating an analyte from a sample in the millisecond time domain.
Accordingly, a need exists for a column with dimensions that are capable of achieving a high rate of selective separation in the millisecond time domain.
Among the several aspects of the invention therefore, is provided a method of loading a microcolumn comprising an active layer and an inert layer, the active layer being capable of separating an analyte from a sample, the method comprising introducing the active layer into the microcolumn such that the active layer is capable of separating the analyte from the sample within the millisecond time domain, and introducing the inert layer.
Another aspect of the invention is provided a microcolumn for separating an analyte from a sample, the microcolumn comprising an active layer capable of separating the analyte from the sample in the millisecond time domain, and an inert layer.
In yet a further aspect is provided a method for separating an analyte from a sample in the millisecond time domain, the method comprising applying the sample to the microcolumn described herein.